c) Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) has been t
ID: 195934 • Letter: C
Question
c) Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) has been the most recent breakthrough discovery in bioengineering that enables scientists to edit DNA. Because you have studied biology in this course, you have volunteered at your niece's Middle School Science Club to monitor a student debate about CRISPR. The students will be watching the following video before the discussion: https://www youtube.com/watch?time continue-252&v-2pp17E4E-08; and you need to be prepared in case there are any questions. Please research and write an answer to each of the following questions: What is "CRISPR"? What role does Cas9 play in the CRISPR process? How does the CRISPR-Cas9 system snip and replace any DNA sequence? What are the potential benefits and drawbacks of gene editing? Include specific examples. Do you believe that the inherent risks of modifying animal DNA is worth the rewards? Explain. Do you believe that it is ethical to genetically engineer humans and/or animals? Explain.Explanation / Answer
Answer:
1. CRISPR-Cas9 is a unique technology that enables geneticists and medical researchers to edit parts of the genome by removing, adding or altering sections of the DNA sequence.
2. The CRISPR-Cas9 system consists of two key molecules that introduce a change (mutation) into the DNA. These are: Cas9 and guide RNA (gRNA)
Cas9 is an enzyme which acts as a pair of ‘molecular scissors’ that can cut the two strands of DNA at a specific location in the genome so that bits of DNA can then be added or removed.
3. The gRNA is a piece of RNA which consists of a small piece of pre-designed RNA sequence (about 20 bases long) located within a longer RNA scaffold. The scaffold part binds to DNA and the pre-designed sequence ‘guides’ Cas9 to the right part of the genome. This makes sure that the Cas9 enzyme cuts at the right point in the genome.
The guide RNA is designed to find and bind to a specific sequence in the DNA. The guide RNA has RNA bases that are complementary to those of the target DNA sequence in the genome. This means that, at least in theory, the guide RNA will only bind to the target sequence and no other regions of the genome.
The Cas9 follows the guide RNA to the same location in the DNA sequence and makes a cut across both strands of the DNA. At this stage the cell recognises that the DNA is damaged and tries to repair it.
Scientists can use the DNA repair machinery to introduce changes to one or more genes? in the genome of a cell of interest.
4. Benefits of gene editing:
Arguably, the most important advantages of CRISPR/Cas9 over other genome editing technologies is its simplicity and efficiency.
Since it can be applied directly in embryo, CRISPR/Cas9 reduces the time required to modify target genes compared to gene targeting technologies based on the use of embryonic stem (ES) cells. Improved bioinformatics tools - to identify the most appropriate sequences to design guide RNAs - and optimization of the experimental conditions enabled very robust procedures which guarantee successful introduction of the desired mutation.
Drawbacks:
The molecular mechanism exploited to insert DNA fragments (e.g. cDNAs) is mediated by DNA repair machinery activated by the double strand break introduced by Cas9. Since the scope of the DNA repair system is not to integrate DNA fragments in the genome, targeted alleles often carry additional modifications, such as deletions, partial or multiple integrations of the targeting vector, and even duplications.
Secondary unwanted mutational events at the target locus plague standard ES cell based projects as well, and researchers have learned how to avoid generating mice carrying passenger mutations. To identify the correct recombination events in ES cells, most laboratories use a combination of positive and negative selection procedures and validation procedures aimed at detecting additional mutations at the target site.
When performing the CRISPR/Cas9 procedure directly on embryos, on the other hand, it is impossible to select for the desired event, greatly limiting the possibility to identify the desired allele. Moreover, the mosaicism observed in founder mice generated using the CRISPR/Cas9 approach makes the identification of unwanted genomic modifications at the target site very challenging.
(Since there are multiple questions, the first four questions have been answered according to the rules of Chegg)
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